Crean, St-John; (2001) Surface associated exported proteins from Staphylococcus aureus, their role in cytokine modulation and possible relevance to bone infection. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

With the emergence of methicillin resistant Staphylococcus aureus (MRSA), the search is on to find alternative therapeutic targets. The work in the thesis is based upon an hypothesis that surface associated exported proteins (SAEP), found within the easily removable surface associated layer on S. aureus, are important in interacting with host cells, resulting in pathological processes central to diseases such as osteomyelitis. These include: a) platelet thrombosis b) endothelial cell activation c) pro-inflammatory cytokine stimulation d) cytokine release modulation. SAEP from S. aureus FRI 326, in the presence of nonaggregating concentrations of ADP, induced platelet aggregation. Further work identified that S. aureus strain 16 was capable of activating an endothelial cell line to release pro- inflammatory cytokine IL-6 and the chemokine IL-8. This activity was independent of IL-1, cyclooxygenase and lipoxygenase enzymes and utilised both tyrosine kinase and p38 signal transduction pathways. SAEP from S. aureus strains 16 and FRI 326 induced both IL-6 and IL-8 release from endothelial, epithelial, peripheral blood mononuclear cells and primary osteoblasts. Attempts to identify specific stimulatory proteins yielded two candidates, one of 50kDa and another of 60kDa. Activity was independent of LPS contamination and utilised protein kinase C, tyrosine kinase and p38 signal transduction pathways. SAEP from S. aureus FRI 326 appeared to inhibit LPS induced cytokine release from peripheral blood mononuclear cells. Initial sequencing data suggested a novel protein. Further sequence studies following re-purification, determined the identity of this protein to be sphingomyelinase (βk-haemolysin). Sphingomyelinase stimulated the release of pro-inflammatory cytokines from Mono-Mac-6, epithelial, endothelial and primary osteoblast cells. In contrast, it was cytocidal to peripheral blood mononuclear cells. This work has demonstrated that SAEP from S. aureus may be capable of a) promoting platelet aggregation, b) activating endothelial cells, c) stimulating the release of pro-inflammatory cytokines and d) modulating cytokine release from a variety of cell types. Each of these processes plays an important role in the establishment of bone infection such as osteomyelitis and as such offer potential therapeutic targets.

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